Mechanism of Insoluble Aggregate Formation in a Reconstituted Solution of Spray-Dried Protein Powder.
Pharm Res
; 40(10): 2355-2370, 2023 Oct.
Article
en En
| MEDLINE
| ID: mdl-37131104
BACKGROUND: Spray-drying is considered a promising alternative drying method to lyophilization (freeze-drying) for therapeutic proteins. Particle counts in reconstituted solutions of dried solid dosage forms of biologic drug products are closely monitored to ensure product quality. We found that high levels of particles formed after reconstitution of protein powders that had been spray-dried under suboptimal conditions. METHODS: Visible and subvisible particles were evaluated. Soluble proteins in solution before spray-drying and in the reconstituted solution of spray-dried powder were analyzed for their monomer content levels and melting temperatures. Insoluble particles were collected and analyzed by Fourier transform infrared microscopy (FTIR), and further analyzed with hydrogen-deuterium exchange (HDX). RESULTS: Particles observed after reconstitution were shown not to be undissolved excipients. FTIR confirmed their identity as proteinaceous in nature. These particles were therefore considered to be insoluble protein aggregates, and HDX was applied to investigate the mechanism underlying aggregate formation. Heavy-chain complementarity-determining region 1 (CDR-1) in the aggregates showed significant protection by HDX, suggesting CDR-1 was critical for aggregate formation. In contrast, various regions became more conformationally dynamic globally, suggesting the aggregates have lost protein structural integrity and partially unfolded after spray-drying. DISCUSSION: The spray-drying process could have disrupted the higher-order structure of proteins and exposed the hydrophobic residues in CDR-1 of the heavy chain, contributing to the formation of aggregate through hydrophobic interactions upon reconstitution of spray-dried powder. These results can contribute to efforts to design spray-dry resilient protein constructs and improve the robustness of the spray-drying process.
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Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Proteínas
/
Microscopía
Idioma:
En
Revista:
Pharm Res
Año:
2023
Tipo del documento:
Article
País de afiliación:
Estados Unidos
Pais de publicación:
Estados Unidos